Friday, December 12, 2014

Drug
resistant infections have become a major problem in modern day medicine. Humans
develop drugs to fight an organism. The organism responds by developing counter
measures to block the drugs, rendering them useless. This back and forth
adaptation causes an arms race between humans and organisms, as each tries to
gain an advantage and become the victor. Arsenical drugs work by taking advantage of an
amino-purine transporter in Trypanosoma
brucei to get the drugs into the cell (1). Melarsoprol B is a
melaminophenyl arsenical drug whose mechanism of action is not fully
understood. It is known to get into the cells using P2, an amino-purine
transporter also called TbAT1, and inbibits glycolytic enzymes,
phosphogluconate dehydrogenase, and trypanothione reductase (2). Trypanosoma brucei is a protozoan parasite that is the causative
agent of African Sleeping Sickness in humans, as well as other diseases in
various species. Trypanosoma brucei
has developed a way to prevent arsenical drugs from killing the parasite. One
such mechanism makes use of the adenosine transport system (3). T. brucei has a mutation in the adenosine transporter that prevents
the cells to take up both exogenous adenosine and arsenical drugs. The
evolution of drug resistance has started an arms race between humans and
pathogens that shows no signs of being resolved.

Trypanosoma brucei comes in three different varieties, which can infect both animals
and humans. T. brucei brucei causes
Nagana in cattle (4). The two varieties that cause
African Sleeping Sickness in humans are T.
brucei gambiense and T. brucei
rhodsiense. T. brucei gambiense
is responsible for roughly 98% of T.
brucei infections and found in more than 24 countries in West and central
Africa (4). T. brucei gambiense, causes a chronic infection, in which symptoms
do not manifest for months to years after infection. T. brucei rhodsiense, on the other hand, makes up the last two
percent of T. brucei infections and
is limited to thirteen countries in Eastern and Southern Africa. This type of
infection is more acute and symptoms develop weeks to months after infection(4). African Sleeping Sickness,
or African Trypanosomiasis, symptoms can be broken down into two stages: stage
one symptoms and stage two symptoms. Stage one symptoms are characterized by
fever, headache, weakness, itching, and joint pain. Treating African Sleeping
Sickness after the manifestation of first stage symptoms is effective, however
it is difficult to diagnose from first stage symptoms (5). Second stage symptoms are
more severe; symptoms include convulsions, confusion, and violent behavior. One
other interestingsymptom of African
Trypanosomiasis is that patients are often unable to sleep at night, yet become
overwhelmed by sleepiness during the day (5). Arsenical drugs are used as
second stage treatments (4). Since these drugs are more
toxic and can cross the blood brain barrier, they are reserved for more severe
symptoms(4). Some common examples of
arsenical drugs are Melarsoprol and Trypursamide(6). These drugs are extremely
toxic and are only used when the infection has spread to the central nervous
system. Because of the late detection of Trypanosomiasis, the parasite has
established a strong hold on the host. The strongly established growth in the
host makes it so that the drugs create a selective environment where the
resistant strains are able to recolonize the host once the competition is
eliminated.African sleeping sickness is
difficult to treat since some strains of T.
brucei have developed mechanisms for resistance. These strains have a
mutation in the gene that encodes an adenosine transporter which prevents cells
from pumping arsenicals into the cell. To test the role of adenosine
transporters in arsenical resistance, genes for TbAT1 from both susceptible and
resistant strains of T. brucei brucei
were cloned into purine auxotrophic Saccharomyces
cerevisiae. The yeast, like T.
brucei, are unable to make their own adenosine and therefore cannot survive
unless they pump adenosine into the cell (3). This experiment makes use of
the characteristic nature of yeast, which do not normally take up exogenous
adenosine. After yeast cells were transformed with the gene for TbAT1 from T. brucei brucei, they were grown on
plates containing adenosine, which the cells normally are unable to take up,
and adenine, which cells readily take up. Next, the amount of adenosine and
adenine transported into the cell was measured. Resistant strains and control
yeast were not able to take up adenosine and were not able to grow when plated
on 150 μM adenosine concentrations.

There are two types of adenosine
transporters, called P1 and P2(3), which differ in specificity.
P1 transporters are specific for adenosine and inosine. P2 transporters are
specific for adenosine, adenine, and melaminophenyl arsenicals (3). In this study, they tested
the inhibition of adenine transport in the presence of inosine and other
arsenicals. Adenosine transport was not inhibited in the presence of inosine,
indicating that inosine was not the substrate of the adenosine transport pump. Melarsoprol,
melarsen oxide, and isometamidium are all substrates for the adenosine
transporter and in the presence of each of these arsenical drugs, adenosine
transport into the cell is inhibited, indicating that the substrate of the pump
was adenosine and arsenical drugs not adenosine and inosine. Therefore, the
pump exhibits P2 activity, not P1 activity (3). While the genes used were
from T. brucei brucei, the results
can be translated to other subspecies of T.
brucei.

The war between humans and pathogens rages on,
as each species fights for life. Parasitologists study these parasites so they
can understand how the parasites develop resistance to drugs. By understanding
the mechanisms of drug resistance, scientists can develop other drugs that will
fight these infections. Just when they have figured out the mechanism of drug
resistance and start treating the infection, the parasite finds a new mechanism
of drug resistance. The parasites must respond to the development of new drugs
by building up their arsenal and developing novel ways to resist being killed
by a new drug. They invoke many different strategies to prevent pumping the
drugs into the cell or by creating new enzymes to break down drug components.
This fight between humans to kill pathogens and parasites to fight drugs has
led to an arms race between humans and parasites that continues to escalate
with no end in sight.

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